Porcine somatic cell nuclear transfer (SCNT) is currently inefficient, as 1–3.95% of reconstructed embryos survive to term; inadequate or erroneous epigenetic reprogramming of the specialized donor somatic nucleus could be a primary reason. Therefore, a locus-specific analysis of DNA methylation dynamics in embryogenesis and the DNA methylation status of gametes and donor cells used for SCNT were conducted in the following developmentally important gene loci: POU5F1, NANOG, SOX2, H19, IGF2, IGF2R, XIST; and the retrotransposon LINE-1. There were significant epigenetic differences between the gametes and the somatic donor cells. Three gamete-specific differentially methylated regions (DMRs) in POU5F1, XIST, and LINE-1 were identified. A delayed demethylation process at POU5F1 and LINE-1 loci occurred after three successive cleavages, compared to the in vitro fertilized (IVF) embryos. Although cloned embryos could undergo de-methylation and re-methylation dynamics at the DMRs of imprinted genes (H19,IGF2R, and XIST), the re-methylation process was compromised, unlike in fertilized embryos. LINE-1 loci are widely dispersed across the whole genome, and LINE-1 DMR might be a potential porcine nuclear reprogramming epi-marker. Data from observations in our present and previous studies, and two published articles were pooled to produce a schematic diagram of locus-specific, DNA methylation dynamics of cloned and IVF embryos during porcine early embryogenesis. This also indicated aberrant DNA methylation reprogramming events, including inadequate DNA demethylation and insufficient re-methylation in cloned embryos. Further research should focus on mechanisms underlying demethylation during the early cleavage of embryos and de novo DNA methylation at the blastocyst stage.

中文翻译：

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克隆和体外受精猪胚胎中 DNA 甲基化模式的位点特异性分析

猪体细胞核移植 (SCNT) 目前效率低下，因为 1-3.95% 的重建胚胎存活到足月；专门供体体细胞核的表观遗传重编程不充分或错误可能是主要原因。因此，对胚胎发生中 DNA 甲基化动力学以及用于 SCNT 的配子和供体细胞的 DNA 甲基化状态的位点特异性分析在以下发育重要基因位点进行：POU5F1、NANOG、SOX2、H19、IGF2、IGF2R、XIST；和反转录转座子LINE-1。配子和体细胞供体细胞之间存在显着的表观遗传差异。鉴定了 POU5F1、XIST 和 LINE-1 中的三个配子特异性差异甲基化区域 (DMR)。POU5F1 和 LINE-1 基因座的延迟去甲基化过程发生在三个连续切割后，与体外受精（IVF）胚胎相比。尽管克隆胚胎可以在印迹基因（H19、IGF2R 和 XIST）的 DMR 处经历去甲基化和再甲基化动力学，但与受精胚胎不同，再甲基化过程受到了损害。LINE-1 基因座广泛分布在整个基因组中，LINE-1 DMR 可能是一种潜在的猪核重编程表观标记。我们目前和以前的研究中的观察数据以及两篇已发表的文章被汇集在一起​​，以生成猪早期胚胎发生过程中克隆胚胎和 IVF 胚胎的位点特异性 DNA 甲基化动力学示意图。这也表明异常的 DNA 甲基化重编程事件，包括克隆胚胎中 DNA 去甲基化不足和再甲基化不足。